Accumulation and aggregation of mutant proteins are common traits across different neurodegenerative disorders, such as the polyglutamine expansion disorder Huntington's disease (HD). A recently emerging theme is that if mutant protein accumulation is eliminated, symptomatic progression not only halts but also recovers. For example, in an inducible model of Huntington's disease, loss of mutant protein accumulation in symptomatic animals led to complete reversion of the disease-like symptoms. Therefore, if we can accelerate the clearance of a disease-causing mutant protein, there exists the tantalizing possibility of recovery from disease. But how do cells clear these mutant proteins? And how does clearance of the mutant proteins lead to recovery from symptoms? To gain insight into these questions we have run Affymetrix gene arrays on stably transfected cell lines that carry mutant huntingtin protein. The comparison of the different genetic profiles revealed surprisingly robust changes in pathways indicating lysosome-mediated degradation and vesicular trafficking. These two areas are little explored in Huntington's disease and polyglutamine diseases in general, and is thus a rich source of questions. In this proposal we will therefore systematically test the following hypotheses: 1) Lysosome-mediated degradation has a significant impact on the degradation of mutant huntingtin proteins; and 2) Aggregation leads to reversible deficits in vesicular trafficking. Using a combination of biochemical and genetic techniques we also propose to identify regulators of protein aggregation and clearance using a functional cell-based assay: a stable cell line that conditionally expresses mutant proteins fused to variants ol GFP. In sum, during this grant period we will reveal targets that directly alter the level of mutant proteins in a cell, elucidate the basic degradation pathways crucial for handling these difficult proteins, and examine how deficits in protein degradation alters vesicular trafficking in the cell. [unreadable] [unreadable]